There is increasing evidence that graft-specific antibodies (Abs) play a dominant role in xenograft rejection, and a critical role in allograft rejection. While anti-graft IgM antibodies are probably the most critical initiators of hyperacute rejection of xenografts and allografts, the roles of anti-graft IgG antibodies in transplantation rejection have not been well characterized. In this proposal we have focussed on anti-galactose-c_1,3- galactose (Gal) Abs that play a dominant role in xenograft rejection. We have generated a series of 53 anti- Gal producing hybridomas, and have extensively characterized these anti-Gal monoclonal Abs (mAbs) in vitro. We have also developed a novel model system using double knockout mice, deficient in the galactosyltransferase (GT) and recombination-activating (Rag-l) genes, as recipients of Gal-expressing heart grafts in conjunction with passively transfused anti-Gal IgG monoclonal Abs (mAbs). We will use this model to investigate the mechanisms by which anti-Gal mAbs elicit graft rejection or induce accommodation. Graft accommodation is defined as the survival of grafted organs in the presence of anti-graft Abs and normal complement levels. The specific aims of this proposal are to define the mechanisms of anti-Gal IgGl-mediated rejection and graft accommodation. We have recently demonstrated that an anti-Gal IgG1 antibody has the ability to induce a form of very acute rejection (VAR) that is distinct from the hyperacute rejection mediated by anti-Gal IgG3. Specifically, we will investigate the contribution of complement and NK cell-mediated inflammatory processes to the pathogenicity of anti-Gal IgGl. We will also define the in vivo mechanisms of IgG-mediated graft accommodation. We observed that the administration of purified anti-Gal IgG mAbs at doses that do not elicit xenograft rejection is not a neutral event, but induces graft accommodation. Thus, xenografts pretreated with low doses of anti-Gal IgG1 acquire resistance and do not succumb to rejection when challenged with high doses of anti-Gal IgG1 mAbs. We propose to characterize the in vivo conditions that favor the development of graft accommodation and to define the molecular mechanisms of anti-Gal IgG-induced accommodation. We anticipate that these studies will lead to an improved understanding of the roles of antibodies in graft rejection and accommodation, and the development of new strategies to improve graft function in humans.